The present invention relates to novel compounds and medical methods of treatment of inflammation, atherosclerosis, restenosis, and immune disorders especially those associated with lymphocyte or monocyte accumulation such as arthritis and transplant rejection. More particularly, the present invention concerns the use of 2-phenyl benzimidazole derivatives.
Migration of leukocytes from blood vessels into diseased tissues is important to the initiation of normal disease-fighting inflammatory responses. But this process, known as leukocyte recruitment, is also involved in the onset and progression of debilitating and life-threatening inflammatory and autoimmune diseases. The pathology of these diseases results from the attack of the body""s immune system defenses on normal tissues. Thus, blocking leukocyte recruitment to target tissues in inflammatory and autoimmune disease would be a highly effective therapeutic intervention. The leukocyte cell classes that participate in cellular immune responses include lymphocytes, monocytes, neutrophils, eosinophils and basophils. In many cases, lymphocytes are the leukocyte class that initiates, coordinates, and maintains chronic inflammatory responses, and thus are generally the most important class of cells to block from entering inflammatory sites. Lymphocytes attract monocytes to the site, which, collectively with lymphocytes, are responsible for much of the actual tissue damage that occurs in inflammatory disease. Infiltration of lymphocytes and/or monocytes is responsible for a wide range of chronic, autoimmune diseases, and also organ transplant rejection. These diseases include, but are not limited to, rheumatoid arthritis, atherosclerosis, psoriasis, chronic contact dermatitis, inflammatory bowel disease, multiple sclerosis, sarcoidosis, idiopathic pulmonary fibrosis, dermatomyositis, skin pemphigoid and related diseases, (e.g., pemphigus vulgaris, p. foliacious, p. erythematosis), glomerulonephritides, vasculitides, hepatitis, diabetes, allograft rejection, and graft-versus-host disease.
This process, by which leukocytes leave the bloodstream and accumulate at inflammatory sites, and initiate disease, takes place in at least three distinct steps which have been described as (1) rolling, (2) activation/firm adhesion and (3) transendothelial migration [Springer, T. A., Nature 346:425-433 (1990); Lawrence and Springer, Cell 65:859-873 (1991); Butcher, E. C., Cell 67:1033-1036 (1991)]. The second step is mediated at a molecular level by chemoattractant receptors. Chemoattractant receptors on the surface of leukocytes bind chemoattractant cytokines secreted by cells at the site of damage or infection. Receptor binding activates leukocytes, increases the adhesiveness of the adhesion molecules that mediate transendothelial migration, and promotes directed migration of the cells toward the source of the chemoattractant cytokine.
A recent discovery is the existence of a large family ( greater than 20 members) of structurally homologous chemoattractant cytokines, approximately 8 to 10 kD in size. These molecules share the ability to stimulate directed cell migration (chemotaxis) and have been collectively called xe2x80x9cchemokinesxe2x80x9d, a contraction of chemotactic cytokines. Each chemokine contains four cysteine residues (C) and two internal disulfide bonds. Chemokines can be grouped into two subfamilies, based on whether the two amino terminal cysteine residues are immediately adjacent (Cxe2x80x94C family) or separated by one amino acid (Cxe2x80x94Xxe2x80x94C family). These differences correlate with the organization of the two subfamilies into separate gene clusters. Within each gene cluster, the chemokines typically show sequence similarities between 25 to 60%.
The chemokines of the Cxe2x80x94Xxe2x80x94C subfamily, such as interleukin-8, are produced by a wide range of cell types and act predominantly on neutrophils as mediators of acute inflammation. Chemokines of the Cxe2x80x94C subfamily are also produced by a wide variety of cell types. These molecules act predominantly on subsets of mononuclear inflammatory cells. Currently there are at least six Cxe2x80x94C chemokines with known chemotactic activity for human monocytes and/or T cells, including MCP-1, MCP-2, MCP-3, MIP-1xcex1, MIP-1xcex2, and RANTES. This suggests there may be a high degree of redundancy in chemoattractant pathways. In addition, most Cxe2x80x94C chemokines are chemotactic for more than one cell type. For examples, RANTES (regulated on activation, normal T cell expressed and secreted) acts on memory CD4+ T cells, eosinophils, and monocytes. Monocyte chemoattractant protein-1 (MCP-1), another Cxe2x80x94C chemokine, acts on monocytes, activated xe2x80x9cmemoryxe2x80x9d T cells and on basophils. MCP-1 is also a potent secretogogue of inflammatory mediators for monocytes and basophils.
Five Cxe2x80x94C chemokine receptors have recently been characterized (CKR1-5 or CCR1-CCR5), and all of these belong to the seven transmembrane spanning G protein-coupled receptor family. Each of these receptors mediates the binding and signaling of more than one chemokine. For example, the CCR1 receptor binds both MIP-1xcex1 and RANTES. There are 2 receptors which bind MCP-1, CCR2 (with alternately spliced forms, 2A and 2B) and CCR4. CCR2 is also known to mediate binding and signaling of MCP-3. The CCR4 receptor binds and signals, in addition to MCP-1, with RANTES and MIP-1xcex1. Which of these is responsible for the MCP-1 mediated recruitment of monocytes and T cells is not known.
In agreement with the observation that lymphocyte emigration into inflammatory sites is usually accompanied by emigration of monocytes, MCP-1 is expressed at sites of antigen challenge and autoimmune disease. However, analyses of human inflammatory lesions with antibodies to other chemokines show RANTES, MIP-1xcex1, MIP-1 and MCP-3 to be present as well. Injection of MCP-1 into skin sites in mice provokes only a mild monocytic infiltrate or no infiltrate at all (Ernst, C. A. et al., J. Immunol. 152:3541-3544, 1994). Whether these results reflect redundant and complex recruitment pathways has not been resolved. MCP-1 and MCP-3 may play a role in allergic hypersensitivity disease. This is suggested by the observation that MCP-1 lacking the amino terminal glutamic acid loses the ability to stimulate basophil mediator release and acquires activity as an eosinophil chemoattractant.
Chemokines of both subfamilies may bind to heparan sulfate proteoglycans on the endothelial cell surface, and may function principally to stimulate haptotaxis of leukocytes that attach to cytokine-activated endothelium through induced adhesion molecules. Additionally, MCP-1 has been reported to selectively activate the xcex21 integrin family of leukocyte adhesion molecule, suggesting a role in leukocyte interactions with the extracellular matrix. Hence, MCP-1 may not only trigger the initial arrest and adhesion of monocytes and T cells, but may also act to guide their migration in extravascular space.
Chemoattractants appear to be required for transendothelial migration in vitro and in vivo and can induce all steps required for transmigration in vivo. Injection of neutrophil chemoattractants into skin or muscle leads to robust emigration of neutrophils from the vasculature and accumulation at the injection site (Colditz, 1991). Pretreatment of neutrophils with pertussis toxin inhibits emigration into inflammatory sites (Spangrude, et al., 1985; Nourshargh and Williams, 1990). Moreover, MAb to IL-8 markedly inhibits neutrophil emigration in inflammation (Sekido et al., 1993).
Neutrophil chemoattractants injected into the same skin site hours apart will stimulate neutrophil accumulation the first time but not the second time, whereas a second injection into a distant site will stimulate accumulation at that site. This desensitization occurs for homologous chemoattractants only (Colditz, 1991) or those that interact with the same receptor. Thus, chemoattractants can act on and homologously desensitize a cell type that is localized in tissue.
Chemoattractants impart directionality to leukocyte migration. By contrast with intradermal injection, intravascular injection of IL-8 does not lead to emigration (Hechtman et al., 1991). Cytokine-stimulated endothelial monolayers grown on filters secrete IL-8 into the underlying collagen layer. Neutrophils added to the apical compartment emigrate into the basilar compartment, but not when the IL-8 gradient is disrupted by addition of IL-8 to the apical compartment (Huber et al., 1991).
The endothelium may present chemoattractants to leukocytes in a functionally relevant way, as well as providing a permeability barrier that stabilizes the chemoattractant gradient. Since leukocytes, responding to specific antigen or inflammatory signals in tissue, may signal emigration of further leukocytes into the site, a chemoattractant was sought in material secreted by mitogen-stimulated mononuclear cells (Carr et al., 1994). Purification to homogeneity guided by a transendothelial lymphocyte chemotaxis assay revealed that monocyte chemoattractant protein 1 (MCP-1), previously thought to be solely a monocyte chemoattractant, is a major lymphocyte chemoattractant. An activated subset of memory lymphocytes respond to MCP-1. In the same assay, lymphocytes respond to RANTES and MIP-1xcex1 but less so than to MCP-1 (Cxe2x80x94C chemokines) and not at all to IL-8 or IP-10 (Cxe2x80x94Xxe2x80x94C chemokines). This physiologically relevant assay suggests that Cxe2x80x94C chemokines tend to attract both monocytes and lymphocytes. In agreement with the observation that lymphocyte emigration into inflammatory sites is accompanied by emigration of monocytes, MCP-1 is abundantly expressed at sites of antigen challenge and autoimmune disease (Miller and Krangel, 1992) and, together with other chemokines, is an excellent candidate to provide the step 2 signal required to activate integrin adhesiveness and emigration of lymphocytes in vivo. (Traffic Signals for Lymphocyte Recirculation and Leukocyte Emigration: The Multistep Paradigm; Springer, 1994, Cell 76: 301-314).
We have surprisingly found that 2-phenyl benzimidazole derivates are MCP-1 receptor antagonists and are capable of inhibiting the binding of MCP-1 to its receptor. Surprisingly, the compounds block T cell migration in vitro, and more surprisingly still, have dramatic effects on the recruitment of inflammatory cells in multiple models of inflammatory diseases. Thus, these compounds are useful as agents for the treatment of inflammatory disease, especially those associated with lymphocyte and/or monocyte accumulation, such as arthritis, atherosclerosis and transplant rejection. In addition, these compounds can be used in the treatment of allergic hypersensitivity disorders such as asthma and allergic rhinitis characterized by basophil activation and eosinophil recruitment, as well as for the treatment of restenosis and chronic or acute immune disorders.
Accordingly, a first embodiment of the present invention provides a method of treatment of chronic or acute inflammatory disease, atherosclerosis, restenosis, chronic or acute immune disorders, and transplant rejection in mammals in need thereof comprising administering to such mammal an effective amount of a 2-phenyl benzimidazole of Formula I or a pharmaceutically acceptable salt thereof: 
wherein A is N or CH;
W, X, Y, and Z can be independently Cxe2x80x94R2, Cxe2x80x94R3, Cxe2x80x94R4, Cxe2x80x94R5, or N,
no more than two of W, X, Y, and Z can be N in any one structure,
R2, R3, R4, and R5 can be independently
H,
C1-20 alkyl,
halogen,
nitro,
xe2x80x94SO2NR8R9,
alkoxy of from 1-4 carbon atoms,
xe2x80x94S(O)pR where p is an integer of from 0 to 2,
xe2x80x94(CH2)mOR,
xe2x80x94(CH2)mCOOR,
xe2x80x94(CH2)mNR8R9,
xe2x80x94(CH2)mCONR8R9,
xe2x80x94(CH2)mCOR,
xe2x80x94CF3,
xe2x80x94benzyl, or
phenyl wherein benzyl or phenyl is optionally substituted with one or two substituents each independently selected from alkyl, halogen, hydrogen, hydroxy, or alkoxy;
m is an integer of from 0 to 4,
R is hydrogen, lower alkyl of from 1-4 carbon atoms, aryl of from 6-10 carbon atoms or benzyl;
when X and Y are substituted by alkyl, they can be joined to form a ring fused at X and Y;
R1 is H, lower alkyl of from 1-4 carbon atoms, or xe2x80x94(CH2)mxe2x80x94Ph;
Rxe2x80x22 is:
H,
C1-20 alkyl,
halogen,
nitro,
xe2x80x94SO2NR8R9,
alkoxy of from 1-4 carbon atoms,
xe2x80x94S(O)pR wherein p is an integer of from 0 to 2,
xe2x80x94(CH2)mOR1xe2x80x94CH2COOR,
xe2x80x94(CH2)mNR8R9,
xe2x80x94(CH2)mCONR8R9,
xe2x80x94(CH2)mCOR, or
xe2x80x94CF3;
R6 is hydrogen, alkyl of from 1 to 6 carbon atoms or R7;
R7 is (CH2)n NR10R11;
n is an integer from 2 to 6;
R8 and R9 can be independently hydrogen, lower alkyl of from 1-4 carbon atoms or can be taken together to form a ring of from 3-8 atoms having up to one additional heteroatom as O, S, SO2, or Nxe2x80x94R12;
R10 and R11 can independently be lower alkyl, xe2x80x94(CH2)mPh, unsubstituted or substituted with up to three R2 substituents, or
R10 and R11 can be taken together to form a ring of from 3 to 8 atoms which may contain oxygen or NR12;
R12 is
hydrogen,
lower alkyl,
xe2x80x94(CH2)tPh, where Ph is phenyl unsubstituted or substituted with up to three R2 substituents;
t is an integer of from 0 to 2;
or a pharmaceutically acceptable salt thereof.
A still further and second embodiment of the present invention is a method of treatment of atherosclerosis in mammals in need thereof comprising administering to such mammal an effective amount of a compound selected from the group consisting of: a compound of Formula I in combination with one or more agents selected from the group consisting of:
(a) ACAT inhibitor;
(b) HMG-CoA reductase inhibitor;
(c) Lipid regulator; and
(d) Bile acid sequestrant;
or a pharmaceutically acceptable salt thereof.
Also, the invention is directed to inhibiting the binding of MCP-1 by utilizing an effective inhibiting amount of a compound of Formula I.
Also, the invention is directed to the novel compositions of Formula I.
Finally, the present invention is directed to a pharmaceutical composition for administering an effective amount of a compound of Formula I in unit dosage form in the treatment methods mentioned above.